Congenital cardiac malformation is the most frequent birth defect and it contributes to advanced heart failure in the pediatric and adult population. An enhanced understanding of the transcriptional networks that direct cardiac progenitors during heart development will have important applications related to cell based therapies and the treatment of congenital heart disease. Furthermore, a number of parallel transcriptional pathways or networks have been proposed for the generation and regeneration of tissues such as the heart. For these reasons, we predict that the definition of the transcriptional regulatory mechanisms of cardiac progenitor cells in the developing heart will enhance our understanding of cardiogenesis, congenital heart disease and myocardial regeneration. We have recently utilized transgenic technology to isolate and characterize cardiac progenitor cells from the developing mouse heart. We have identified Etsrp71 as a potential downstream target of Nkx2-5 and generated Etsrp71 deficient embryos that are nonviable and have perturbed cardiovascular development. Our principle hypothesis is that Nkx2-5 regulated networks direct discrete stages of cardiac morphogenesis. We further predict that the Etsrp71 is a direct downstream target of Nkx2-5 and is a master regulator of the endocardial/endothelial lineages. In these proposed studies, we take an innovative approach to examine these hypotheses and employ emerging technologies to define the role of the Nkx2-5 - Etsrp71 cascade and cardiovascular development. To examine these hypotheses, we will address the following specific aims: 1) To define the Nkx2.5 transcriptional regulatory cascades in cardiac progenitor cells isolated from the developing murine heart. 2) To define the functional role of Etsrp71 during embryogenesis. 3) To determine whether Etsrp71 is a master regulator of the endocardial/endothelial lineage. These studies will enhance our understanding of the role of transcriptional networks and signaling pathways that mechanistically regulate cardiac progenitor cell (CPC) populations to acquire a cardiovascular fate during development. In addition, the results of these studies will serve as a platform for therapeutic strategies directed towards congenital heart disease and advanced heart failure. PUBLIC HEALTH RELEVANCE: Congenital heart disease is common and deadly. An increased understanding of the networks that direct cardiac progenitors and stem cells during heart development will have important therapeutic applications for the treatment of congenital heart disease. As the regulation of these progenitor cells are largely unknown, this proposal will decipher the molecular pathways that govern cardiac progenitors, which may ultimately serve as a platform to enhance the regenerative process in patients with cardiac malformations and advanced heart failure.